Publications

4477

ABSTRACT In the past 10 years, the number of endosymbionts described within the bacterial order Rickettsiales has constantly grown. Since 2006, 18 novel Rickettsiales genera inhabiting protists, such as ciliates and amoebae, have been described. In this work, we characterize two novel bacterial endosymbionts from Paramecium collected near Bloomington, IN. Both endosymbiotic species inhabit the cytoplasm of the same host. The Gram-negative bacterium “ Candidatus Bealeia paramacronuclearis” occurs in clumps and is frequently associated with the host macronucleus. With its electron-dense cytoplasm and a distinct halo surrounding the cell, it is easily distinguishable from the second smaller symbiont, “ Candidatus Fokinia cryptica,” whose cytoplasm is electron lucid, lacks a halo, and is always surrounded by a symbiontophorous vacuole. For molecular characterization, the small-subunit rRNA genes were sequenced and used for taxonomic assignment as well as the design of species-specific oligonucleotide probes. Phylogenetic analyses revealed that “ Candidatus Bealeia paramacronuclearis” clusters with the so-called “basal” Rickettsiales , and “ Candidatus Fokinia cryptica” belongs to “ Candidatus Midichloriaceae.” We obtained tree topologies showing a separation of Rickettsiales into at least two groups: one represented by the families Rickettsiaceae , Anaplasmataceae , and “ Candidatus Midichloriaceae” (RAM clade), and the other represented by “basal Rickettsiales ,” including “ Candidatus Bealeia paramacronuclearis.” Therefore, and in accordance with recent publications, we propose to limit the order Rickettsiales to the RAM clade and to raise “basal Rickettsiales ” to an independent order, Holosporales ord. nov., inside Alphaproteobacteria , which presently includes four family-level clades. Additionally, we define the family “ Candidatus Hepatincolaceae” and redefine the family Holosporaceae . IMPORTANCE In this paper, we provide the characterization of two novel bacterial symbionts inhabiting the same Paramecium host (Ciliophora, Alveolata). Both symbionts belong to “traditional” Rickettsiales , one representing a new species of the genus “ Candidatus Fokinia” (“ Candidatus Midichloriaceae”), and the other representing a new genus of a “basal” Rickettsiales . According to newly characterized sequences and to a critical revision of recent literature, we propose a taxonomic reorganization of “traditional” Rickettsiales that we split into two orders: Rickettsiales sensu stricto and Holosporales ord. nov. This work represents a critical revision, including new records of a group of symbionts frequently occurring in protists and whose biodiversity is still largely underestimated.

AbstractGut-associated microbiota of ants include Rhizobiales bacteria with affiliation to the genus Bartonella. These bacteria may enable the ants to fix atmospheric nitrogen, but no genomes have been sequenced yet to test the hypothesis. Sequence reads from a member of the Rhizobiales were identified in the data collected in a genome project of the ant Harpegnathos saltator. We present an analysis of the closed 1.86 Mb genome of the ant-associated bacterium, for which we suggest the species name Candidatus Tokpelaia hoelldoblerii. A phylogenetic analysis reveals a relationship to Bartonella and Brucella, which infect mammals. Novel gene acquisitions include a gene for a putative extracellular protein of more than 6,000 amino acids secreted by the type I secretion system, which may be involved in attachment to the gut epithelium. No genes for nitrogen fixation could be identified, but genes for a multi-subunit urease protein complex are present in the genome. The urease genes are also present in Brucella, which has a fecal-oral transmission pathway, but not in Bartonella, which use blood-borne transmission pathways. We hypothesize that the gain and loss of the urease function is related to transmission strategies and lifestyle changes in the host-associated members of the Rhizobiales.

Abstract“Candidatus Liberibacter asiaticus” (CLas), a non-culturable α-proteobacterium, is associated with citrus Huanglongbing (HLB, yellow shoot disease) currently threatening citrus production worldwide. Here, the whole genome sequence of CLas strain A4 from Guangdong of China was analyzed. Five copies of nrdB, encoding β-subunit of ribonucleotide reductase (RNR), a critical enzyme involving bacterial proliferation, were found. Three nrdB copies were in long form (nrdBL, 1,059 bp) and two were in short form (nrdBS, 378 bp). nrdBS shared >99% identity to 3′ end of nrdBL and had no active site. Sequences of CLas nrdB genes formed a distinct monophyletic lineage among eubacteria. To make use of the high copy number feature, a nrdB-based primer set RNRf/RNRr was designed and evaluated using real-time PCR with 262 HLB samples collected from China and USA. Compared to the current standard primer set HLBas/HLBr derived from the 16S rRNA gene, RNRf/RNRr had Ct value reductions of 1.68 (SYBR Green PCR) and 1.77 (TaqMan PCR), thus increasing the detection sensitivity three-fold. Meanwhile, RNRf/RNRr was more than twice the stability of primer set LJ900f/LJ900r derived from multi-copy prophage. The nrdB-based PCR thereby provides a sensitive and reliable CLas detection with broad application, especially for the early diagnosis of HLB.

Abstract Ni.tro.so.te'nu.is. N.L. adj. nitrosus, full of natron; here intended to mean nitrous; L. masc. adj. tenuis slender; N.L. masc. n. Candidatus Nitrosotenuis a slender nitrite producer. Candidatus Nitrosotenuis: a slender nitrite producer. Candidatus Nitrosotenuis is a genus of Thaumarchaeota that can be found widely distributed in soils, freshwater, hot springs, subsurface, and activated sludge. They may be rods or spheres and may or may not have flagella. Similar to other known and described Thaumarchaeota , Ca . Nitrosotenuis are aerobic chemolithautotrophs that use energy gained from the oxidation of ammonia to nitrite to fix carbon via the modified 3‐hydroxypropionate/4‐hydroxybutyrate carbon fixation pathway. At this time, no pure culture of Ca . Nitrosotenuis exists, but some of the six available enrichments have only a few rare contaminants. It remains to be demonstrated whether the remaining bacterial contaminants provide essential compounds to Ca . Nitrosotenuis. Enrichments of Ca . Nitrosotenuis are intolerant to high salinity (>0.3%), although they are phylogenetically placed within the group I.1a Thaumarchaeota ( Ca . Nitrosopumilales), which includes taxa that are widely distributed in the ocean. DNA G + C content (mol%) : 41.0–42.2. Type species : Candidatus Nitrosotenuis uzonensis . Lebedeva, Hatzenpichler, Pelletier, Schuster, Hauzmayer, Bulaev, Grigor'eva, Galushko, Schmid, Palatinszky, Le Paslier, Daims and Wagner 2013, 8. Taxonomic and Nomenclature Notes According to the List of Prokaryotic names with Standing in Nomenclature (LPSN), the taxonomic status of the genus Candidatus Nitrosotenuis is: preferred name (not correct name) (last update, February 2025) * . LPSN classification: Archaea / Thermoproteati / Thermoproteota / Nitrososphaeria / Nitrosopumilales / Candidatus Nitrosotenuaceae / Candidatus Nitrosotenuis The genus Candidatus Nitrosotenuis can also be recovered in the Genome Taxonomy Database (GTDB) as g__Nitrosotenuis (version v220) ** . GTDB classification: d__Archaea / p__Thermoproteota / c__Nitrososphaeria / o__Nitrososphaerales / f__Nitrosopumilaceae / g__Nitrosotenuis * Meier‐Kolthoff et al. ( 2022 ). Nucleic Acids Res , 50 , D801 – D807 ; DOI: 10.1093/nar/gkab902 ** Parks et al. ( 2022 ). Nucleic Acids Res , 50 , D785 – D794 ; DOI: 10.1093/nar/gkab776

Abstract Ni.tro.so.te.nu.a.ce'ae. N.L. adj nitrosus nitrous; L. adj. tenuis slender; ‐aceae ending to denote a family; N.L. fem. pl. n. Nitrosotenuaceae the Nitrosotenuis family. Candidatus Nitrosotenuaceae: A family of slender nitrite producers Candidatus Nitrosotenuaceae is a family of Thaumarchaeota that consists of a single genus, Ca . Nitrosotenuis, which can be found widely distributed in soils, freshwater, hot springs, the subsurface, and activated sludge. They may be rods or spheres and may or may not have flagella. Similar to other known and described Thaumarchaeota , Ca . Nitrosotenuaceae are aerobic chemolithautotrophs that use energy gained from the oxidation of ammonia to nitrite to fix carbon via modified 3‐hydroxypropionate/4‐hydroxybutyrate carbon fixation pathway. At this time, no pure culture of Ca . Nitrosotenuaceae exists, but some of the six available enrichments with members of the genus are almost pure. It remains to be demonstrated whether the remaining bacterial contaminants provide essential compounds to Ca . Nitrosotenuaceae. Enrichments of Ca . Nitrosotenuaceae are intolerant to high salinity (>0.3%), although they are phylogenetically related to the Group I.1a Thaumarchaeota ( Ca . Nitrosopumilaceae), which includes taxa that are widely distributed in the ocean. Taxonomic and Nomenclature Notes According to the List of Prokaryotic names with Standing in Nomenclature (LPSN), the taxonomic status of the family Candidatus Nitrosotenuaceae is: preferred name (not correct name) (last update, February 2025) * . LPSN classification: Archaea / Thermoproteati / Thermoproteota / Nitrososphaeria / Nitrosopumilales / Candidatus Nitrosotenuaceae The family Candidatus Nitrosotenuaceae can also be recovered in the Genome Taxonomy Database (GTDB) as f__Nitrosopumilaceae (version v220) ** . GTDB classification: d__Archaea / p__Thermoproteota / c__Nitrososphaeria / o__Nitrososphaerales / f__Nitrosopumilaceae * Meier‐Kolthoff et al. ( 2022 ). Nucleic Acids Res , 50 , D801 – D807 ; DOI: 10.1093/nar/gkab902 ** Parks et al. ( 2022 ). Nucleic Acids Res , 50 , D785 – D794 ; DOI: 10.1093/nar/gkab776

Abstract Cel.lu.lo.si.ly.ti.ca.ce'ae. N.L. neut. n. Cellulosilyticum a bacterial genus; N.L. fem. pl. n. Cellulosilyticaceae the Cellulosilyticum family. Slightly curved rod‐shaped cells . Gram‐positive cell‐wall structure but stain Gram‐negative. Terminal spores formed. Mesophilic, optimum growth temperature is approximately 40°C. Obligate anaerobic . No microaerophilic or aerobic growth occurs. Produce acetate, ethanol, and/or formate as the major end products, but not butyric or propionic acid. Hydrolyze cellulose and xylan and ferment cellobiose, but not ferment lactate and pyruvate. The predominant fatty acids are C 16:0 (27.1 %), C 14:0 (9.2 %), and iso‐C 16:0 (6.4%). Phylogenetically distant from the family Lachnospriaceae (Clostridum cluster XIVb), a member of the phylum Firmicutes, with less than 87.7% 16S rRNA similarity. Thus far, only one genus Cellulosilyticum is described. DNA G + C content (mol%) : 33–36. Type genus : Cellulosilyticum Cai and Dong 2010, 848 VP . Taxonomic and Nomenclature Notes According to the List of Prokaryotic names with Standing in Nomenclature (LPSN), the taxonomic status of the family Cellulosilyticaceae is: synonym (last update, February 2025) * . Correct name: Lachnospiraceae LPSN classification: Bacteria / Bacillati / Bacillota / Clostridia / Eubacteriales / Lachnospiraceae The family Cellulosilyticaceae can also be recovered in the Genome Taxonomy Database (GTDB) as f__Cellulosilyticaceae (version v220) ** . GTDB classification: d__Bacteria / p__Bacillota_A / c__Clostridia / o__Lachnospirales / f__Cellulosilyticaceae * Meier‐Kolthoff et al. ( 2022 ). Nucleic Acids Res , 50 , D801 – D807 ; DOI: 10.1093/nar/gkab902 ** Parks et al. ( 2022 ). Nucleic Acids Res , 50 , D785 – D794 ; DOI: 10.1093/nar/gkab776